CLEARED TISSUE LIGHTSHEET

Cleared Tissue LightSheet (CTLS) is a large field light sheet

microscope designed to image whole organs at high speed.

CTLS creates a focused sheet with a narrow waist for better

optical sectioning, then uses a spatial light modulator (SLM)

to rapidly shift the waist of the sheet along the axis of

propagation. A dual excitation setup allows imaging from the

right and left sides of the specimen for optimal light sheet

projection throughout. Piezoelectric stages move the

specimen in x, y, and z with sub-micron resolution. The result

is clear: a 1 cm3 volume can be imaged at up to 1µm x 1µm x

3µm (XYZ) resolution, and a cleared mouse brain can be

imaged in as little as 1.5 hours.

High Speed High Resolution Imaging of Cleared Tissue and Whole Organs

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The Difference is ClearLight sheet microscopy is a powerful technique for imaging large specimens by taking full advantage of emerging tissue clearing

methods. The chemistry behind these techniques has advanced to where we can easily penetrate 1, 5 even 10mm into a specimen with

a focused sheet of light. In combination with a macro zoom microscope using high NA large field of view lenses, Cleared Tissue

LightSheet can image large field sizes with high resolution in short periods of time.

XY

XZ

No Tiling 20 Tiles

How Tiling Works

+ + =

The optical sectioning ability of a light sheet is dependent on the thickness of the waist of the focused beam used to create the sheet.

The thickness of the waist is directly proportional to the beam length. A thinner waist is generally required for better optical

sectioning, but the thinner the waist the shorter the usable length of the beam. Imaging large cleared specimens with a light sheet

requires a beam with a long waist matched to the large field of view. The long waist has a correspondingly high thickness, and the

result is often poor optical sectioning.

To dramatically improve on this limitation, CTLS uses a spatial light modulator to create a sharply focused beam with a thin waist much

shorter than the detector's field of view. The beam waist is tiled along the axis of propagation and the camera is synchronized to

capture one image per tile. The optimal region of each capture is selected and stitched together forming a continuously optimized

image. The resulting data has excellent axial resolution compared to a non-translated focused beam.

Benefits of CTLS• Excellent resolution from the macro objectives

• Excellent optical sectioning from the thin waist of the tiled light sheet

• Low photobleaching via light sheet as compared to confocal

• Ultrafast acquisition from large-format macro objectives, large-format sCMOS camera, and digital focus via SLM eliminating the

need to move optical elements

• Fully automated hardware control, image acquisition and data stitching via SlideBook software

CTLS acquisition is extremely flexible, from

ultrafast capture with a 20µm light sheet

(left) to high-resolution capture with a 3µm

light sheet shifted 20 times and the

resulting 20 sections of best focus tiled to

one best-focus image (right). Images

compare lateral (XY) and axial (XZ) views

of the ventral Tegmental Nuclear (VTN)

group of the mouse cleared with PEGASOS

(Jing et al. (2018). Tissue clearing of both

hard and soft tissue organs with the

PEGASOS method. Cell Research).

Sample courtesy of Dr. Hu Zhao (Texas A&M

University).

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FeaturesDUAL LEFT AND RIGHT ILLUMINATION

HIGH NA / LARGE FIELD OF VIEW /

LONG WORKING DISTANCE OBJECTIVES

PIEZO STAGES DRIVING MULTIPLE

SAMPLE CHAMBERS

OPTICAL ZOOM PRESCAN

LASERSTACK LASER COMBINER

GPU-OPTIMIZED SLIDEBOOK SOFTWARE

LARGE DATA SOLUTIONS

Allows for optimal sheet penetration across wide specimens and avoidance of opaque

structures that may be present on one side but not the other.

Macro lenses at 1x/0.25NA and 1.5x/0.37NA can image through any cleared organ with

excellent resolution.

Sub-micron resolution stages allow precise positioning of the specimen in sample

chambers sized to fit the biology.

CTLS includes a motorized optical zoom to automatically zoom out and create a 2D

map of the entire specimen. This map serves as virtual eyepieces allowing inspection of

the entire specimen at higher magnification and identification of regions of interest for

zoomed-in high resolution imaging in 3D.

Fiber-coupled laser combiner allows up to six lasers covering the entire visible

spectrum at multiple power levels.

SlideBook directs all hardware synchronization and data capture, creating 3D datasets

at over 1TB ready for analysis and rendering.

Available DDN® unified storage systems allow direct acquisition and analysis without

time-consuming file transfers for 200TB to over 1PB.

SlideBook allows scientists to focus on investigation rather than

instrumentation, controlling every aspect of CTLS from hardware

configuration to image acquisition, data reconstruction, image

processing and 3D visualization. With over 20 years of active

development in collaboration with researchers worldwide,

SlideBook is intuitive yet powerful with features from visual

imaging ROI selection to automated SLM pattern generation and

axial chromatic aberration correction. SlideBook SLD files can be

accessed via any application supporting Bio-Formats OME,

allowing seamless collaboration in any workflow.

Advanced FeaturesAXIAL CHROMATIC ABERRATION

CORRECTION

AUTOMATIC REDUCTION OF

SHADOWING & STRIPING EFFECTS

Leveraging the SLM, SlideBook creates patterns specific to each laser wavelength to

correct for axial chromatic aberration.

Light sheets can be subject to distortion as they encounter objects that have not been

cleared, resulting in shadows or striping in the image plane. SlideBook uses the SLM to

create a patterned light sheet that interrogates the specimen from 3 different angles to

mitigate (and in some instances completely eliminate) these artifacts.

BUILT BY SCIENTISTS FOR SCIENTISTS. Intelligent Imaging Innovations (3i) designs

and manufactures cutting edge live cell and intravital microscopy imaging platforms

driven by 64-bit SlideBook software. 3i was established in 1995 by a group of scientists

whose wide range of research activities includes cell biology, immunology, neuroscience

and computer science. Our collective aim is to provide advanced multi-dimensional

microscopy platforms that are intuitive to use, modular in design, and meet the

evolving needs of investigators in the biological research community.

Printed on 100% post-consumer recycled paper

NORTH AMERICA

Intelligent Imaging Innovations, Inc.

3509 Ringsby Court

Denver, CO 80216 USA

+1 303 607 9429

Fax +1 303 607 9430

info@intelligent-imaging.com

www.intelligent-imaging.com

EUROPE

Intelligent Imaging Innovations, GmbH

Königsallee 9-21

D-37081 Göttingen, Germany

+49 (0) 551 508 39 266

Fax +49 (0) 551 508 39 268

Intelligent Imaging Innovations, Ltd

7 Westbourne Studios

242 Acklam Rd

London, UK W10 5JJ

+44 (0) 2037 440031

3ieurope@intelligent-imaging.com

Specifications1 µm x 1 µm x 3 µm

1x/0.25NA, 1.5x/0.37NA

< 1 min/mm3

25mm x 25mm x 25mm (XYZ)

Multiple glass specimen chambers optimized for 1x/0.25NA and 1.5x/0.37NA objectives, with specimen

holder. Custom-shaped specimen holders available upon request.

Organic and aqueous clearing solutions

2048x2048 16-bit sCMOS

LaserStack compact modular laser launch with 488nm and 561nm lasers standard. Up to 4 additional

wavelengths upon request.

Piezoelectric with sub-micron resolution in X, Y and Z

SlideBook™ software for acquisition and GPU-accelerated analysis

RESOLUTION

OBJECTIVES

IMAGING SPEED

SAMPLE TRAVEL RANGE

SPECIMEN CHAMBERS

COMPATIBLE CLEARING

METHODS

CAMERA

LASER LINES

XYZ TRANSLATION STAGE

SOFTWARE

Mouse mandible cleared with PEGASOS

showing vasculature and dentin in the

molar teeth reported by endogenous

fluorescence cdh5cre. Sample courtesy

of Dr. Dian Jing and Dr. Yating Li, Sichuan

University, Department of Orthodontics.

Smooth muscle cells in the arteries, veins

and capillaries of mouse brain cleared with

PEGASOS reported by NG2BacDsRed.

Sample courtesy of Dr. Woo-Ping Ge,

University of Texas Southwestern Medical

Center.

Mouse hind paw labelled with Thy-1 YFP

and cleared with PEGASOS. Specimen

courtesy of Dr. Wenjing Luo, Texas A&M

Health Sciences Center.